PROJECT SUMMARY Summary of Parent R01: LMNA mutations accounts for approximately 6% of cases in familial dilated cardiomyopathies (DCM). Compared to other DCM patients, LMNA cardiolaminopathy patients exhibit a severe clinical course, conduction abnormalities, and a high rate of heart failure. LMNA DCM has a complex pathophysiology and it has been hypothesized that this form of cardiac disease is due to defects in function of cardiomyocytes (CMs) as well as defects in non-CM populations such as endothelial cells (ECs). However, the detailed molecular mechanisms of LMNA cardiolaminopathy remain elusive. Accordingly, the parent R01 proposes to generate human induced pluripotent stem cell- derived cardiomyocytes (iPSC-CMs) from LMNA DCM patients as well as matched family controls and healthy unrelated controls. To illuminate molecular mechanisms, we are performing structural, electrophysiological, transcriptomic, and mechanistic analyses using patient-specific as well as genome-edited isogenic iPSC-CMs and iPSC-ECs. We will also perform drug screening studies on these cell types to target dysregulated signaling pathways and identify targeting compounds. Collectively, these studies are anticipated to uncover mechanistic insights of LMNA cardiolaminopathy, a major cause of DCM, and may help identify novel therapeutic candidates that can target CMs and ECs, two key cell populations. Proposed Supplement: In the proposed Research Supplement, we will extend the parent R01 to include two critically related sets of experiments. First, we will expand the cohort of DCM patients and healthy control to include 20 additional individuals of African American ethnicity, which will lay the groundwork for exploring population-specific effects of DCM and allow systematic comparison of population differences in the proposed drug screening effort. Secondly, we will take advantages of latest advances in single-cell RNA sequencing (scRNA-seq) and characterize DCM iPSC-CMs and iPSC-ECs at a single cell level and identify disease mechanisms free from confounding effects of cellular heterogeneity. The proposed experiments will be performed by, and will enable the training of, Mr. Damon Williams, a recent baccalaureate graduate who is a member of a group underrepresented in biomedical research (African Americans).